Approximately 6.5 million Americans are diagnosed with a cutaneous chronic wound each year, costing the healthcare system in the United States over 20 billion dollars annually. Often these chronic wounds are found in geriatric individuals, up to 85% of patients with chronic wounds are over 65 years old, or in patients with concurrent diseases such as diabetes, obesity, malnutrition, or substance abuse. Similarly, the failure to properly heal wounds is a major problem for the Veteran population as presently there are over 9 million Veterans over the age of 65, or about 38% of the total Veteran population. Often these patients are subjected to a litany of various treatments, frequently without mechanistic justification, and they still lack adequate resolution of their wounds. Therefore, a better understanding of specific causes of chronic wounds and a better menu of treatment options is needed to address this unmet need. Once any type of chronic wound appears, whether it is a diabetic ulcer, decubitus ulcer, or venous stasis ulcer, one of the primary limiting factors in wound closure is the inability of epidermal cells to re-epithelialize the wound bed. Chronic wounds remain arrested in the inflammatory phase with little to no migration of keratinocytes across the wound bed. Recent data have demonstrated that an evolutionarily-conserved mechanism, called the Integrated Stress Response (ISR), is crucial for normal re-epithelialization in human skin. The ISR consists of four distinct but related protein kinases are activated by various types of environmental stress leading to the phosphorylation of the same target protein, eukaryotic initiation factor 2 on its alpha subunit (eIF2?~P). The consequences of eIF2?~P include a global inhibition of protein translation, which conserves vital cellular energy stores, and the selective translation of genes associated with responding to the stress. Importantly, it was recently demonstrated that the ISR is activated in differentiating keratinocytes in normal skin. Furthermore, keratinocytes deficient for one of the eIF2?~P protein kinases (GCN2, which phosphorylates eIF2?~P during epidermal differentiation) form abnormal, dyskeratotic skin lacking a normal barrier function. If GCN2 expression is knocked-out in human keratinocytes (an exon 12 deletion using CRISPR/CAS9), keratinocytes are deficient in epithelial sheet migration without affecting individual cell motility. These data led to the hypothesis that GCN2 is a critical component of the cutaneous wounding response, and that manipulation of the ISR via pharmaceutical agents will improve wound healing. This last point is particularly intriguing, because several drugs that regulate the ISR (either positively or negatively) are already in human clinical trials for unrelated diseases which could accelerate bringing any drugs that showed promise rapidly into human trials. The initial studies into the role of the ISR on wound healing in this proposal use model systems including two- dimensional keratinocyte growth in vitro as well as three-dimensional organ skin systems and human clinical studies in vivo. In vitro studies will define the mechanism of GCN2 activation following wounding of the keratinocyte cell culture, determine which function in keratinocyte sheet migration is compromised following GCN2 inactivation, and analyze which genes and signaling pathways are activated by enhanced protein translation using a combination of polysome profiling and RNA-seq analyses. In vivo studies will determine the utility of ISR-specific activators/inhibitors on cutaneous wound healing, determine if the inactivation of normal ISR in geriatric patients alters normal wound healing, and examine pathological biopsies from chronic wounds to determine is the ISR is inactivated during the progression of this disease. The management of chronic wounds, as well as the potential to accelerate resolution of acute cutaneous injuries, is of critical importance to Veterans, both as quality of life issues for Veteran patients and cost-effectiveness of medical care for the VA system. It is the objective of this proposal to identify specific therapeutic or mechanistic pathways that can alleviate the consequences of chronic wounds.